This application is a 371 of PCT/JP98/03730, filed Aug. 24, 1998, which claims the priority of Japan JP9-241699, filed Aug. 25, 1997
The present invention relates to novel neuropeptides and more specifically to peptides obtained from the venoms of the solitary wasps Anoplius samariensis and Batozonellus maculifrons, and analogous peptides thereof.
Since solitary wasp species such as thread-waisted wasps and pompilid wasps paralyze other insects with their venom, it is believed that the venom contain substances that act on the nervous system. In fact, peptides called mastoparans have been isolated from a certain kind of solitary wasp, and these peptides are used as useful compounds for the study of information transmission system of the living body.
On the other hand, various compounds have been isolated in the studies on spider toxins, and it has been confirmed that their effect of paralyzing nerves is based on their inhibitory action on glutamate receptors. Glutamic acid is known to play an important role as an excitatory neurotransmitter mediated by glutamate receptors in the central nervous system (e.g. brain, spinal cord) of mammals, and the synapses of insects and crustaceans. And further, it is known that excess glutamic acid causes over-excitation of nerve cells resulting in death of the nerve cells. Therefore, enhancement of the function of glutamate receptors and conversely temporary blocking of their function are believed to provide therapeutic benefits on cerebral nerve diseases associated with glutamic acid.
Accordingly, the object of the present invention is to find, in the venom of the solitary wasps, substances that act on the glutamate receptor or substances that promote or inhibit the release of glutamic acid, and to apply them to medical therapeutic purposes.
In order to achieve the above object, the present inventors have isolated a peptide (1) represented by SEQ ID NO: 1:
H-Arg-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2xe2x80x83xe2x80x83(1)
and a peptide (97) represented by SEQ ID NO: 2:
H-Arg-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2xe2x80x83xe2x80x83(97)
from the venom sac extracts of solitary wasps A. samariensis and B. maculifrons, on the basis of the ability to enhance or block the neurotransmission through the neuron synapses of lobster""s walking leg.
Furthermore, peptides related to the above peptides (1) and (97) were synthesized by the conventional solid phase method and their structure-activity relationship was examined. As a result, it was found that the peptides represented by the amino acid sequence formula (A):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Rxe2x80x83xe2x80x83(A)
wherein:
R is basically NH2, which also may be -Leu-NH2, -Leu-Ser-NH2, -Leu-Ser-Lys-NH2, -Leu-Ser-Arg-NH2, -Leu-Ser-Lys-Leu-NH2, (SEQ ID NO:3) or -Leu-Ser-Arg-Leu-NH2 (SEQ ID NO:4),
X1 and X3 are basic amino acid residues;
X2, X4 and X6 are aliphatic amino acid residues; and
X7 is an aromatic amino acid residue, can show the above-mentioned activity.
The basic amino acid residues X1 and X3 include arginine or lysine residue, the aliphatic amino acid residues X2, X4 and X6 include leucine or isoleucine residue, and the aromatic amino acid residue X7 includes phenylalanine, tyrosine, or tryptophan residue.
Accordingly, the present invention provides, as a neuro-peptide, a peptide represented by the amino acid sequence formula (A):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Rxe2x80x83xe2x80x83(A)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp; and
R represents xe2x80x94NH2, -Leu-NH2, -Leu-Ser-NH2, -Leu-Ser-Lys-NH2, -Leu-Ser-Arg-NH2, -Leu-Ser-Lys-Leu-NH2, (SEQ ID NO:3) or -Leu-Ser-Arg-Leu-NH2 (SEQ ID NO:4);
and a salt thereof.
As an embodiment, the present invention provides a peptide represented by the amino acid sequence formula (B):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-NH2xe2x80x83xe2x80x83(B)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
As another embodiment, the present invention provides a peptide represented by the amino acid sequence formula (C):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Leu-NH2(C)xe2x80x83xe2x80x83(SEQ ID NO:200)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
As a further embodiment, the present invention provides a peptide represented by the amino acid sequence formula (D):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Leu-Ser-NH2(D)xe2x80x83xe2x80x83(SEQ ID NO:199)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
As still another embodiment, the present invention provides a peptide represented by the amino acid sequence formula (E):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Leu-Ser-Lys-NH2(E)xe2x80x83xe2x80x83(SEQ ID NO:5)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
As still another embodiment, the present invention provides a peptide represented by the amino acid sequence formula (F):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Leu-Ser-Arg-NH2(F)xe2x80x83xe2x80x83(SEQ ID NO:6)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
As still another embodiment, the present invention provides a peptide represented by the amino acid sequence formula (G):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Leu-Ser-Lys-Leu-NH2(G)xe2x80x83xe2x80x83(SEQ ID NO:7)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
As still a further embodiment, the present invention provides a peptide represented by the amino acid sequence formula (H):
H-X1-X2-X3-X4-Gly-X6-X7-Asp-Gln-Leu-Ser-Arg-Leu-NH2(H)xe2x80x83xe2x80x83(SEQ ID NO:8)
wherein:
each of X1 and X3 independently represents Arg or Lys, each of X2, X4 and X6 independently represents Ile or Leu, and X7 represents Phe, Tyr or Trp;
and a salt thereof.
Among the above peptides provided by the present invention, the most preferred embodiment is a peptide represented by the amino acid sequence formula (1):
H-Arg-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2(1)xe2x80x83xe2x80x83(SEQ ID NO:1)
and a salt thereof, or a peptide represented by the amino acid sequence formula (97):
H-Arg-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2(97)xe2x80x83xe2x80x83(SEQ ID NO:2)
and a salt thereof.
As another embodiment, the present invention provides a pharmaceutical composition containing the peptide represented by the amino acid sequence formula (A) or a salt thereof for treatment of glutamate receptor related diseases.
As still another embodiment, the present invention provides use of the peptide represented by the amino acid sequence formula (A) or a salt thereof for treatment of glutamate receptor related diseases.
As still a further embodiment, the present invention provides a method for treatment of glutamate receptor related diseases which composing administering an effective amount of the peptide represented by the amino acid sequence formula (A) or a salt thereof.
In the present specification and in each of the above formulae, Gly stands for a glycine residue, Asp stands for an aspartic acid residue, Gln stands for a glutamine residue, Arg stands for an arginine residue, Lys stands for a lysine residue, Ile stands for an isoleucine residue, Leu stands for a leucine residue, Phe stands for a phenylalanine residue, Tyr stands for a tyrosine residue, Trp stands for a tryptophan residue, and Ser stands for a serine residue. Amino acid sequences are indicated by the triplet code of IUPAC.
The peptides provided by the present invention include the following peptides:
H-Arg-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (1) (SEQ ID NO:1)
H-Arg-Ile-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (2) (SEQ ID NO:9)
H-Arg-Ile-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (3) (SEQ ID NO:10)
H-Arg-Ile-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (4) (SEQ ID NO:11)
H-Arg-Ile-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (5) (SEQ ID NO:12)
H-Arg-Ile-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (6) (SEQ ID NO:13)
H-Arg-Ile-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (7) (SEQ ID NO:14)
H-Arg-Ile-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (8) (SEQ ID NO:15)
H-Arg-Ile-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (9) (SEQ ID NO:16)
H-Arg-Ile-Lys-Leu-Gly-leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (10) (SEQ ID NO:17)
H-Arg-Ile-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (11) (SEQ ID NO:18)
H-Arg-Ile-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (12) (SEQ ID NO:19)
H-Arg-Ile-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (13) (SEQ ID NO:20)
H-Arg-Ile-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (14) (SEQ ID NO:21)
H-Arg-Ile-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (15) (SEQ ID NO:22)
H-Arg-Ile-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (16) (SEQ ID NO:23)
H-Arg-Ile-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (17) (SEQ ID NO:24)
H-Arg-Ile-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (18) (SEQ ID NO:25)
H-Arg-Ile-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (19) (SEQ ID NO:26)
H-Arg-Ile-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (20) (SEQ ID NO:27)
H-Arg-Ile-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (21) (SEQ ID NO:28)
H-Arg-Ile-Arg-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (22) (SEQ ID NO:29)
H-Arg-Ile-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (23) (SEQ ID NO:30)
H-Arg-Ile-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (24) (SEQ ID NO:31)
H-Arg-Leu-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (25) (SEQ ID NO:32)
H-Arg-Leu-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (26) (SEQ ID NO:33)
H-Arg-Leu-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (27) (SEQ ID NO:34)
H-Arg-Leu-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (28) (SEQ ID NO:35)
H-Arg-Leu-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (29) (SEQ ID NO:36)
H-Arg-Leu-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (30) (SEQ ID NO:37)
H-Arg-Leu-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (31) (SEQ ID NO:38)
H-Arg-Leu-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (32) (SEQ ID NO:39)
H-Arg-Leu-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (33) (SEQ ID NO:40)
H-Arg-Leu-Lys-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (34) (SEQ ID NO:41)
H-Arg-Leu-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (35) (SEQ ID NO:42)
H-Arg-Leu-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (36) (SEQ ID NO:43)
H-Arg-Leu-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (37) (SEQ ID NO:44)
H-Arg-Leu-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (38) (SEQ ID NO:45)
H-Arg-Leu-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (39) (SEQ ID NO:46)
H-Arg-Leu-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (40) (SEQ ID NO:47)
H-Arg-Leu-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (41) (SEQ ID NO:48)
H-Arg-Leu-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (42) (SEQ ID NO:49)
H-Arg-Leu-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (43) (SEQ ID NO:50)
H-Arg-Leu-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (44) (SEQ ID NO:51)
H-Arg-Leu-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-MH2 (45) (SEQ ID NO:52)
H-Arg-Leu-Arg-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (46) (SEQ ID NO:53)
H-Arg-Leu-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (47) (SEQ ID NO:54)
H-Arg-Leu-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (48) (SEQ ID NO:55)
H-Lys-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (49) (SEQ ID NO:56)
H-Lys-Ile-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (50) (SEQ ID NO:57)
H-Lys-Ile-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (51) (SEQ ID NO:58)
H-Lys-Ile-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (52) (SEQ ID NO:59)
H-Lys-Ile-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (53) (SEQ ID NO:60)
H-Lys-Ile-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (54) (SEQ ID NO:61)
H-Lys-Ile-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (55) (SEQ ID NO:62)
H-Lys-Ile-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (56) (SEQ ID NO:63)
H-Lys-Ile-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (57) (SEQ ID NO:64)
H-Lys-Ile-Lys-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu NH2 (58) (SEQ ID NO:65)
H-Lys-Ile-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (59) (SEQ ID NO:66)
H-Lys-Ile-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (60) (SEQ ID NO:67)
H-Lys-Ile-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (61) (SEQ ID NO:68)
H-Lys-Ile-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (62) (SEQ ID NO:69)
H-Lys-Ile-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (63) (SEQ ID NO:70)
H-Lys-Ile-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (64) (SEQ ID NO:71)
H-Lys-Ile-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (65) (SEQ ID NO:72)
H-Lys-Ile-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (66) (SEQ ID NO:73)
H-Lys-Ile-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (67) (SEQ ID NO:74)
H-Lys-Ile-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (68) (SEQ ID NO:75)
H-Lys-Ile-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (69) (SEQ ID NO:76)
H-Lys-Ile-Arg-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (70) (SEQ ID NO:77)
H-Lys-Ile-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (71) (SEQ ID NO:78)
H-Lys-Ile-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (72) (SEQ ID NO:79)
H-Lys-Leu-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (73) (SEQ ID NO:80)
H-Lys-Leu-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (74) (SEQ ID NO:81)
H-Lys-Leu-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (75) (SEQ ID NO:82)
H-Lys-Leu-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (76) (SEQ ID NO:83)
H-Lys-Leu-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (77) (SEQ ID NO:84)
H-Lys-Leu-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (78) (SEQ ID NO:85)
H-Lys-Leu-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (79) (SEQ ID NO:86)
H-Lys-Leu-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (80) (SEQ ID NO:87)
H-Lys-Leu-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (81) (SEQ ID NO:88)
H-Lys-Leu-Lys-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (82) (SEQ ID NO:89)
H-Lys-Leu-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (83) (SEQ ID NO:90)
H-Lys-Leu-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (84) (SEQ ID NO:91)
H-Lys-Leu-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (85) (SEQ ID NO:92)
H-Lys-Leu-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (86) (SEQ ID NO:93)
H-Lys-Leu-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (87) (SEQ ID NO:94)
H-Lys-Leu-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (88) (SEQ ID NO:95)
H-Lys-Leu-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (89) (SEQ ID NO:96)
H-Lys-Leu-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (90) (SEQ ID NO:97)
H-Lys-Leu-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (91) (SEQ ID NO:98)
H-Lys-Leu-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (92) (SEQ ID NO:99)
H-Lys-Leu-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (93) (SEQ ID NO:100)
H-Lys-Leu-Arg-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (94) (SEQ ID NO:101)
H-Lys-Leu-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (95) (SEQ ID NO:102)
H-Lys-Leu-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Lys-Leu-NH2 (96) (SEQ ID NO:103)
H-Arg-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (97) (SEQ ID NO:104)
H-Arg-Ile-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (98) (SEQ ID NO:105)
H-Arg-Ile-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (99) (SEQ ID NO:106)
H-Arg-Ile-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (100) (SEQ ID NO:107)
H-Arg-Ile-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (101) (SEQ ID NO:108)
H-Arg-Ile-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (102) (SEQ ID NO:109)
H-Arg-Ile-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (103) (SEQ ID NO:110)
H-Arg-Ile-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (104) (SEQ ID NO:111)
H-Arg-Ile-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (105) (SEQ ID NO:112)
H-Arg-Ile-Lys-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (106) (SEQ ID NO:113)
H-Arg-Ile-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (107) (SEQ ID NO:114)
H-Arg-Ile-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (108) (SEQ ID NO:115)
H-Arg-Ile-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (109) (SEQ ID NO:116)
H-Arg-Ile-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (110) (SEQ ID NO:117)
H-Arg-Ile-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (111) (SEQ ID NO:118)
H-Arg-Ile-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (112) (SEQ ID NO:119)
H-Arg-Ile-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (113) (SEQ ID NO:120)
H-Arg-Ile-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (114) (SEQ ID NO:121)
H-Arg-Ile-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (115) (SEQ ID NO:122)
H-Arg-Ile-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (116) (SEQ ID NO:123)
H-Arg-Ile-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (117) (SEQ ID NO:124)
H-Arg-Ile-Arg-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (118) (SEQ ID NO:125)
H-Arg-Ile-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Let-NH2 (119) (SEQ ID NO:126)
H-Arg-Ile-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (120) (SEQ ID NO:127)
H-Arg-Leu-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (121) (SEQ ID NO:128)
H-Arg-Leu-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (122) (SEQ ID NO:129)
H-Arg-Leu-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (123) (SEQ ID NO:130)
H-Arg-Leu-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (124) (SEQ ID NO:131)
H-Arg-Leu-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (125) (SEQ ID NO:132)
H-Arg-Leu-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (126) (SEQ ID NO:133)
H-Arg-Leu-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (127) (SEQ ID NO:134)
H-Arg-Leu-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (128) (SEQ ID NO:135)
H-Arg-Leu-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (129) (SEQ ID NO:136)
H-Arg-Leu-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (130) (SEQ ID NO:137)
H-Arg-Leu-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (131) (SEQ ID NO:138)
H-Arg-Leu-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (132) (SEQ ID NO:139)
H-Arg-Leu-Lys-Leu-Gly-LIe-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (133) (SEQ ID NO:140)
H-Arg-Leu-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (134) (SEQ ID NO:141)
H-Arg-Leu-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (135) (SEQ ID NO:142)
H-Arg-Leu-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (136) (SEQ ID NO:143)
H-Arg-Leu-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (137) (SEQ ID NO:144)
H-Arg-Leu-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (138) (SEQ ID NO:145)
H-Arg-Leu-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (139) (SEQ ID NO:146)
H-Arg-Leu-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (140) (SEQ ID NO:147)
H-Arg-Leu-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (141) (SEQ ID NO:148)
H-Arg-Leu-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (142) (SEQ ID NO:149)
H-Arg-Leu-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (143) (SEQ ID NO:150)
H-Arg-Leu-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (144) (SEQ ID NO:151)
H-Lys-Ile-Lys-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (145) (SEQ ID NO:152)
H-Lys-Ile-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (146) (SEQ ID NO:153)
H-Lys-Ile-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (147) (SEQ ID NO:154)
H-Lys-Ile-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (148) (SEQ ID NO:155)
H-Lys-Ile-Lys-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (149) (SEQ ID NO:156)
H-Lys-Ile-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (150) (SEQ ID NO:157)
H-Lys-Ile-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (151) (SEQ ID NO:158)
H-Lys-Ile-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (152) (SEQ ID NO:159)
H-Lys-Ile-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (153) (SEQ ID NO:160)
H-Lys-Ile-Lys-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (154) (SEQ ID NO:161)
H-Lys-Ile-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (155) (SEQ ID NO:162)
H-Lys-Ile-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (156) (SEQ ID NO:163)
H-Lys-Ile-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (157) (SEQ ID NO:164)
H-Lys-Ile-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (158) (SEQ ID NO:165)
H-Lys-Ile-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (159) (SEQ ID NO:166)
H-Lys-Ile-Arg-Ile-Gly-ILe-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (160) (SEQ ID NO:167)
H-Lys-Ile-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (161) (SEQ ID NO:168)
H-Lys-Ile-Arg-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu NH2 (162) (SEQ ID NO:169)
H-Lys-Ile-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (163) (SEQ ID NO:170)
H-Lys-Ile-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (164) (SEQ ID NO:171)
H-Lys-Ile-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (165) (SEQ ID NO:172)
H-Lys-Ile-Arg-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (166) (SEQ ID NO:173)
H-Lys-Ile-Arg-Leu-Gly-LIe-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (167) (SEQ ID NO:174)
H-Lys-Ile-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (168) (SEQ ID NO:175)
H-Lys-Leu-Lys-ILe-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (169) (SEQ ID NO:176)
H-Lys-Leu-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (170) (SEQ ID NO:177)
H-Lys-Leu-Lys-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (171) (SEQ ID NO:178)
H-Lys-Leu-Lys-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (172) (SEQ ID NO:179)
H-Lys-Leu-Lys-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (173) (SEQ ID NO:180)
H-Lys-Leu-Lys-Ile-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (174) (SEQ ID NO:181)
H-Lys-Leu-Lys-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (175) (SEQ ID NO:182)
H-Lys-Leu-Lys-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (176) (SEQ ID NO:183)
H-Lys-Leu-Lys-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (177) (SEQ ID NO:184)
H-Lys-Leu-Lys-Leu-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (178) (SEQ ID NO:185)
H-Lys-Leu-Lys-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (179) (SEQ ID NO:186)
H-Lys-Leu-Lys-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (180) (SEQ ID NO:187)
H-Lys-Leu-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (181) (SEQ ID NO:188)
H-Lys-Leu-Arg-Ile-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (182) (SEQ ID NO:189)
H-Lys-Leu-Arg-Ile-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (183) (SEQ ID NO:190)
H-Lys-Leu-Arg-Ile-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (184) (SEQ ID NO:191)
H-Lys-Leu-Arg-Ile-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (185) (SEQ ID NO:192)
H-Lys-Leu-Arg-ILe-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (186) (SEQ ID NO:193)
H-Lys-Leu-Arg-Ile-Gly-Ile-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (187) (SEQ ID NO:194)
H-Lys-Leu-Arg-Leu-Gly-Leu-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (188) (SEQ ID NO:195)
H-Lys-Leu-Arg-Leu-Gly-Leu-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (189) (SEQ ID NO:196)
H-Lys-Leu-Arg-Leu-Gly-Leu-Phe-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (190) (SEQ ID NO:197)
H-Lys-Leu-Arg-Leu-Gly-Ile-Tyr-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (191) (SEQ ID NO:198)
H-Lys-Leu-Arg-Leu-Gly-Ile-Trp-Asp-Gln-Leu-Ser-Arg-Leu-NH2 (192) (SEQ ID NO:199)
In order to obtain the peptides of the present invention, for example, venom sac of a solitary wasp containing said peptides may be excised and then homogenized in a water-containing organic solvent, and the resulting homogenate may be centrifuged to obtain a supernatant, which may be purified by a reversed phase chromatography until finally it exhibits a single peak when monitored by UV absorbance at 215 nm, thereby isolating and purifying the desired compound.
The water-containing organic solvent to be used for the homogenization during the above purification process means a mixture of water and an organic solvent that is freely miscible with water, and may be any solvent which permits extraction of the compound of the present invention. Examples include, but not limited to, water-containing acetone, water-containing acetonitrile, and the like. When the resulting supernatant is directly applied to the reversed phase chromatography, it is preferred that about 0.1% of trifluoroacetic acid (hereinafter referred to as TFA) is added to such a water-containing organic solvent.
Since the peptides provided by the present invention consist of at most about 13 amino acids, they can be synthesized by the conventional liquid or solid phase method. Furthermore, since the peptides of the present invention are basic peptides, they can be converted to appropriate acid-added salts according to conventional technique. Such preferred acid-added salts may be obtained by reaction with a pharmaceutically acceptable acid, and said salts include, for example, lactates, acetates, succinates, maleates, fumarates, tartarates, citrates, gluconates, ascorbates, benzoates, methanesulfonates, cinnamates, benzenesulfonates or phosphates, hydrogenphosphates, hydrochlorides, hydrobromides, hydroiodides, sulfamates, sulfates, trifluoroacetates, and the like.
The biological activity of the present peptides can be assayed according to the methods described in Japanese Patent Laid-Open No. 4-198161, in which excitatory postsynaptic potential (EPSP) is generated in the neuromuscular specimen of a lobster""s walking leg, and the effect of the compound given to the nerve fibers on the EPSP is determined.
In general, substances inhibiting EPSP are believed to be glutamate antagonists or inhibitors for the release of glutamic acid, and the substances enhancing EPSP are believed to be glutamate-like agonists or accelerators for the release of glutamic acid.
As described in the Examples below, the peptides of the present invention show a remarkable effect of enhancing EPSP determined by the above assay. Accordingly, the compounds of the present invention are useful for enhancing cerebral neural activities and recovering muscular strength in the treatment of neuromuscular diseases. They are also useful as a tool for the study of neuroscience toward the elucidation of the transmission mechanism.